The long-term goals of the proposed research are to aid in the development of rapid endothelial cell (EC) adherence to smooth muscle cells and formation of a fully functional endothelium within tissue-engineered blood vessels (TEBVs). TEBVs represent a relatively new and promising aproach to treat atherosclerosis. Studies will be performed using our recently optimized human EC-SMC co-culture. The aims of the proposed research are as follows: (1) characterize type of adhesions and integrins involved in the adhesion of ECs; (2) to determine effect of shear stress on EC function when bound to quiescent SMCs or ECM proteins; (3) to determine the relationship between EC integrins and shear stress-induced production of vasoactive molecules and gene regulation. EC integrin blocking experiments will be performed with monoclonal antibodies to determine the major integrin complexes involved with EC adhesion to SMCs. Immunostaining of vinculin and tensin will be performed to determine the major type of EC adhesion (focal vs. fibrillar) to SMCs during static and flow conditions. Vasoactive responses by ECs in co-culture and monoculture to a physiological shear stress will be measured. Finally, the role of EC integrins in response to shear stress will be evaluated by blocking integrin protein expression with siRNA and repeating experiments in Aim 2. Significant differences in the regulation of genes and production of vasoactive molecules found in Aims 2 and 3 will indicate the involvement of these integrins in EC function. Cardiovascular disease, which includes heart diease and stroke, kills more Americans each year than any other disease. Tissue engineered blood vessels can provide doctors a superior alternative material to bypass blocked arteries. This research will advance the knowledge of the cellular interactions within tissue engineered blood vessels. [unreadable] [unreadable] [unreadable]